Plasma cfDNA globality methylation detection method and application

Abstract

The invention discloses a plasma cfDNA globality methylation detection method and application, and relates to the technical field of biological detection. According to the plasma cfDNA globality methylation detection method, methylated exogenous DNA (lambda DNA) containing biotin labels is supplemented to a small amount of cfDNA extracted from plasma, then an enrichment methylation experiment is carried out, after a collected product is subjected to PCR amplification, based on a biotin-streptavidin reaction, streptavidin magnetic beads are adopted for removing the lambda DNA with the biotin labels, a cfDNA amplification library is obtained, and then high-throughput sequencing is conducted. The plasma cfDNA globality methylation detection method is suitable for methylation analysis of a small amount of cfDNA, a kit does not need to be specifically commercialized, the operation is simple and efficient, the sample loss is reduced, the library building efficiency is improved, the detectionsensitivity of MeDIP and other enrichment methylation experiment methods is greatly improved, and the method has wide applicability.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a detection method and application of plasma cfDNA global methylation. Background technique [0002] Plasma free DNA (cfDNA) is free DNA released by cells into peripheral blood. A series of molecular changes that occur in tumor cells, such as methylation modification status, sequence mutation, nucleosome occupancy, etc., can be released from tumors. reflected in cfDNA in blood. Therefore, cfDNA has been extensively studied as a prognostic or predictive marker for cancer detection. Compared with DNA mutations, DNA methylation is more stable and can be continuously measured, because it often occurs in specific regions of DNA called CpG islands; and the diversity of genetic mutations determines the need for cancer diagnosis. A large percentage of the genome is tested to perform a test with sufficient sensitivity. Therefore, detection of cfDNA methylation, as a non-in...

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Problems solved by technology

MeDIP-seq, as a relatively low-cost and widely used methylation detection method, is often used to study the methylome of clinical samples, but usually requires a large amount of sample DNA (initial amount of hundreds of nanometers) gram or even a few micrograms), currently the minimum amount of DNA required to detect plasma cfDNA methylation by MeDIP-seq method is 50ng, if whole blood is used as the starting sample, at least 5mL plasma needs to be separated from 10-20mL whole blood for The extraction of cfDNA requires a large amount of samples, which is not conducive to multiple clinical sampling and storage, which limits the application of cfDNA methylation detection in the fields of tumor diagnosis, drug screening and prognosis monitoring.

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